1. Field of the Invention
The present invention relates to a method of inducing autophagy in a cell, particularly to a method of inducing autophagy in a cell by activating Toll-like receptors.
2. Description of the Prior Art
Graphene and its oxidized form, graphene oxide (GO), have drawn intense attention in recent years for biological and medical applications. The surface of GO contains hydrophilic oxygen-containing functional groups (i.e. hydroxyl, epoxyl and carboxyl tails) on the basal plane and edges, rendering GO amenable to stable dispersion in water and functionalization. These attributes have prompted the use of GO for bioimaging, cellular probing, cellular growth and differentiation, gene and drug delivery and photothermal therapy. These burgeoning applications in biomedicine entail the need to evaluate the in vitro and in vivo safety of GO.
Autophagy is a process that degrades intracellular components in response to stressful conditions (e.g. starvation and infection) and is linked to cellular processes as diverse as cell survival, cell death, pathogen clearance and antigen presentation. Autophagy involves the formation of double-membraned vesicles termed autophagosomes, which sequester cytoplasm and organelles and then fuse with lysosomes to form autolysosomes, thus degrading the contents of the vacuole. Autophagy is negatively controlled by mTOR (mammalian target of rapamycin) complex 1 (mTORC1) and inhibition of mTORC1 kinase activity initiates the formation of autophagosome that comprises a complex consisting of Beclin 1 and other factors. The autophagosome formation also involves the conversion of microtubule-associated protein light chain 3 (LC3-I) to the lipidated form LC3-II, consequently conversion from LC3-I to LC3-II is a common indicator of autophagy.
Toll-like receptors (TLRs) are important receptors for the detection of microbial antigens and subsequent induction of innate immune responses. Among the TLRs, TLR2 recognizes bacterial lipoproteins while TLR3 detects virus-derived dsRNA. TLR4 recognizes lipopolysaccharides (LPS) and TLR5 recognizes bacterial flagellin. TLR7 mediates recognition of viral ssRNA while TLR9 senses unmethylated DNA with CpG motifs derived from bacteria and viruses. Upon engagement with cognate ligands, the TLRs transduce signals by first recruiting adaptor proteins including myeloid differentiating factor 88 (MyD88) and TIR domain-containing adaptor inducing IFN-beta (TRIF), followed by activation of downstream signaling proteins such as TRAF6 and NF-κB, eventually resulting in various cellular responses including secretion of cytokines and interferons (IFNs).
The connection between autophagy and TLRs was discovered in 2007 as it was found that TLRs signaling in macrophages links the autophagy pathway to phagocytosis and TLR4 stimulation enhances the autophagic elimination of phagocytosed mycobacteria in macrophages. Ensuing studies further reported that TLR2, TLR3 and TLR7 play roles in autophagy induction. To date the precise mechanisms regulating the TLRs-elicited autophagy remain to be established although agonists stimulating TLR2, TLR3, TLR4 and TLR7 were shown to trigger autophagy.